DUSP3 is a small dual-specificity protein phosphatase with an unknown physiological function. We report that DUSP3 is strongly expressed in human and mouse monocytes and macrophages and that its ... [more ▼]

DUSP3 is a small dual-specificity protein phosphatase with an unknown physiological function. We report that DUSP3 is strongly expressed in human and mouse monocytes and macrophages and that its deficiency in mice promotes tolerance to lipopolysaccharide (LPS)-induced endotoxin shock and to polymicrobial septic shock following cecal ligation and puncture. By using adoptive transfer experiments, we demonstrate that resistance to endotoxin is macrophage-dependent and transferable and that this protection is associated with a striking increase of M2-like macrophages in DUSP3-/- mice in both the LPS and cecal ligation and puncture models. We show that the altered response of DUSP3-/- mice to sepsis is reflected in decreased TNF production and impaired ERK1/2 activation. Our results demonstrate that DUSP3 plays a key and non-redundant role as a regulator of innate immune responses by mechanisms involving the control of ERK1/2 activation, TNF secretion and macrophage polarization. [less ▲]

Background/Purpose: Immune-mediated inflammatory disorders (IMIDs) share many genetic risk factors. Pleiotropy may exist at different levels and most of the underlying mechanisms are still to be uncovered ... [more ▼]

Background/Purpose: Immune-mediated inflammatory disorders (IMIDs) share many genetic risk factors. Pleiotropy may exist at different levels and most of the underlying mechanisms are still to be uncovered. GWAS have identified hundreds of risk loci for IMIDs but causative genes have been identified in only a handful of cases. Recent fine-mapping efforts indicate that only a minority of risk variants are coding. This suggests that most risk variants will be regulatory hence affecting disease risk via eQTL effects. Methods: To aid in the identification of causative genes for IMIDs, we generated transcriptome information (HT12 arrays) for six blood cell types (CD4, CD8, CD19, CD14, CD15 and platelets) and intestinal biopsies at three anatomical locations (ileum, colon, rectum) for 350 healthy Caucasians. The same individuals were genotyped with SNP arrays interrogating > 700K variants, augmented by imputation from the 1KG project. To detect cis-eQTL we tested variants within 0.5 megabase windows centered on the tested probe. The nominal p-value of the best SNP within a cis-window was Sidak-corrected for the window-specific number of independent tests. The corresponding best, Sidak-corrected p-values for each probe were jointly used to estimate their respective false discovery rate.To identify likely causative genes in GWAS identified risk loci variants and also better understand pleiotropic effects, we (i) developed a method that quantifies the correlation between “disease association pattern” (DAP) and “eQTL association pattern” (EAP) and provides an empirical estimate of its significance, and (ii) evaluated the effect of fitting known risk variants as covariates in the eQTL analysis following Nica et al. (2010). We applied both approaches to celiac disease (CE) and rheumatoid arthritis (RA) and the second one to type one diabetes (T1D), multiple sclerosis (MS), systemic lupus erythematosus (SLE), ankylosing spondylitis (AS) and psoriasis (PSO). Results: We detected > 16000 significant cis-eQTL, with a degree of sharing between cell types ranging from 38 to 90% highlighting the utility of our multi-tissue panel. GWAS variants were drivers of ciseQTL effects across the different tissues in 399 tests (23.6%), mostly in CD4 cells, and pinpointing 64 new gene-disease associations (3.7%). The number of shared loci and shared eQTL were highly correlated (rho=0.66).RA and SLE showed the highest degree of sharing. Conclusions: We identified new potential candidate genes for IMIDs and characterized pleiotropic effects through ciseQTL mapping in GWAS loci. These findings could shed a light on IMIDs pathogenesis and co-occurrence. Latest results will be presented. [less ▲]

Background A limitation of current antiplatelet therapies is their inability to separate thrombotic events from bleeding occurrences. Better understanding of the molecular mechanisms leading to platelet ... [more ▼]

Background A limitation of current antiplatelet therapies is their inability to separate thrombotic events from bleeding occurrences. Better understanding of the molecular mechanisms leading to platelet activation is of importance for the development of improved therapies. Recently, protein tyrosine phosphatases (PTPs) have emerged as critical regulators of platelet function. Methods and Results This is the first report implicating the dual-specificity phosphatase 3 (DUSP3) in platelet signaling and thrombosis. This phosphatase is highly expressed in human and mouse platelets. Platelets from DUSP3-deficient mice displayed a selective impairment of aggregation and granule secretion mediated through the collagen receptor glycoprotein VI (GPVI) and the C-type lectin-like receptor 2 (CLEC-2). DUSP3-deficient mice were more resistant to collagen- and epinephrine-induced thromboembolism, compared to wild-type mice, and showed severely impaired thrombus formation upon ferric chloride-induced carotid artery injury. Intriguingly, bleeding times were not altered in DUSP3-deficient mice. At the molecular level, DUSP3 deficiency impaired Syk tyrosine phosphorylation, subsequently reducing phosphorylation of PLCγ2 and calcium fluxes. To investigate DUSP3 function in human platelets, a novel small-molecule inhibitor of DUSP3 was developed. This compound specifically inhibited collagen and CLEC-2-induced human platelet aggregation, thereby phenocopying the effect of DUSP3 deficiency in murine cells. Conclusions DUSP3 plays a selective and essential role in collagen- and CLEC-2-mediated platelet activation and thrombus formation in vivo. Inhibition of DUSP3 may prove therapeutic for arterial thrombosis. This is the first time a PTP, implicated in platelet signaling, has been targeted with a small-molecule drug. [less ▲]

Genome-wide association studies of the related chronic inflammatory bowel diseases (IBD) known as Crohn's disease and ulcerative colitis have shown strong evidence of association to the major ... [more ▼]

Genome-wide association studies of the related chronic inflammatory bowel diseases (IBD) known as Crohn's disease and ulcerative colitis have shown strong evidence of association to the major histocompatibility complex (MHC). This region encodes a large number of immunological candidates, including the antigen-presenting classical human leukocyte antigen (HLA) molecules. Studies in IBD have indicated that multiple independent associations exist at HLA and non-HLA genes, but they have lacked the statistical power to define the architecture of association and causal alleles. To address this, we performed high-density SNP typing of the MHC in >32,000 individuals with IBD, implicating multiple HLA alleles, with a primary role for HLA-DRB1*01:03 in both Crohn's disease and ulcerative colitis. Noteworthy differences were observed between these diseases, including a predominant role for class II HLA variants and heterozygous advantage observed in ulcerative colitis, suggesting an important role of the adaptive immune response in the colonic environment in the pathogenesis of IBD. [less ▲]

BACKGROUND: The genetic component of Crohn's disease (CD) is well known, with 140 susceptibility loci identified so far. In addition to single nucleotide polymorphisms typically studied in genome-wide ... [more ▼]

BACKGROUND: The genetic component of Crohn's disease (CD) is well known, with 140 susceptibility loci identified so far. In addition to single nucleotide polymorphisms typically studied in genome-wide scans, copy number variation is responsible for a large proportion of human genetic variation. METHODS: We performed a genome-wide search for copy number variants associated with CD using array comparative genomic hybridization. One of the found regions was validated independently through real-time PCR. Serum levels of the found gene were measured in patients and control subjects. RESULTS: We found copy number differences for the C4S and C4L gene variants of complement component C4 in the central major histocompatibility complex region on chromosome 6p21. Specifically, we saw that CD patients tend to have lower C4L and higher C4S copies than control subjects (P = 5.00 x 10 and P = 9.11 x 10), which was independent of known associated classical HLA I and II alleles (P = 7.68 x 10 and P = 6.29 x 10). Although C4 serum levels were not different between patients and control subjects, the relationship between C4 copy number and serum level was different for patients and control subjects with higher copy numbers leading to higher serum concentrations in control subjects, compared with CD patients (P < 0.001). CONCLUSIONS: C4 is part of the classical activation pathway of the complement system, which is important for (auto)immunity. Low C4L or high C4S copy number, and corresponding effects on C4 serum level, could lead to an exaggerated response against infections, possibly leading to (auto)immune disease. [less ▲]

TNF is a master pro-inflammatory cytokine. Activation of TNFR1 by TNF can result in both RIPK1-independent apoptosis and RIPK1 kinase-dependent apoptosis or necroptosis. These cell death outcomes are ... [more ▼]

TNF is a master pro-inflammatory cytokine. Activation of TNFR1 by TNF can result in both RIPK1-independent apoptosis and RIPK1 kinase-dependent apoptosis or necroptosis. These cell death outcomes are regulated by two distinct checkpoints during TNFR1 signaling. TNF-mediated NF-kappaB-dependent induction of pro-survival or anti-apoptotic molecules is a well-known late checkpoint in the pathway, protecting cells from RIPK1-independent death. On the other hand, the molecular mechanism regulating the contribution of RIPK1 to cell death is far less understood. We demonstrate here that the IKK complex phosphorylates RIPK1 at TNFR1 complex I and protects cells from RIPK1 kinase-dependent death, independent of its function in NF-kappaB activation. We provide in vitro and in vivo evidence that inhibition of IKKalpha/IKKbeta or its upstream activators sensitizes cells to death by inducing RIPK1 kinase-dependent apoptosis or necroptosis. We therefore report on an unexpected, NF-kappaB-independent role for the IKK complex in protecting cells from RIPK1-dependent death downstream of TNFR1. [less ▲]

The host immune system plays an critical role in maintaining homeostasis with resident microbial communities, therefore ensuring that the complex symbiotic relationship is maintained. At the same time ... [more ▼]

The host immune system plays an critical role in maintaining homeostasis with resident microbial communities, therefore ensuring that the complex symbiotic relationship is maintained. At the same time, resident microbiota contribute to host nutrition and energy balance and to the development or maintenance of a robust immune system. Dysbiosis of the microbiota is associated with various immunological disorders, including inflammatory bowel diseases (IBD). Both genetic and environmental factors are implicated in this disturbance; however, the relative contributions of these two factors, and the mechanism by which they interact remain unclear. Recently, we started a project that aims to identify molecular components of the hostmicrobiota-connectome by taking advantage of common variation in – on the one hand – the genome, transcriptome and metabolome of the host, and – on the other hand – the composition of its gut microbiota. We will take advantage of the already established CEDAR cohort that provides integrated genetic (SNP genotypes) and transcriptome data (circulating immune cells subset, as well as samples from various anatomical locations in the intestine). We will further enrich the dataset in this cohort with metabolome (plasma), and gut microbiota data (16srRNA sampled at the ileum, colon, and rectum). The CEDAR cohort is composed of healthy individuals and is therefore more suitable to study effect of common risk variants than (IBD) patients, since analysis of samples from patients suffering from active inflammation may only give insight in ongoing patho-physiological processes, that are likely to mask the primum movens events. Next, we will study the overlap between the identified components of the HMC network identified and the ~160 GWAS-identified risk loci for IBD. We anticipate to reveal novel connections between the microbiota and IBD by this integrative “omics” approach, thereby shedding new light on the pathogenesis of IBD. Latest results will be presented with respect to the microbiota composition of from different anatomical locations in the intestine using the V2 and V5-6 regions of the bacterial 16S rRNA. [less ▲]

Lifetime prevalence of inflammatory bowel disease (IBD) is reaching an alarming rate of >1/400 in industrialized societies. Improved understanding of disease pathogenesis is essential to develop more ... [more ▼]

Lifetime prevalence of inflammatory bowel disease (IBD) is reaching an alarming rate of >1/400 in industrialized societies. Improved understanding of disease pathogenesis is essential to develop more effective preventive, diagnostic and therapeutic measures. Genome-wide association studies (GWAS) have identified ~ 160 risk loci contributing to inherited predisposition to IBD, leading to the identification of new perturbed pathways and potential drug targets. Nevertheless, causative genes and variants remain unknown for the vast majority of risk loci. GWAS loci are likely to be regulatory and therefore alter expression levels of other genes. We hypothesize that if an IBD associated SNP is an expression quantitative loci (eQTL)-the " disease-association pattern " (DAP) should mirror the " eQTL association pattern " (EAP) of the causative gene if looking in the right target tissue(s). With this premise, our project aims to detect causative genes implicated in IBD's susceptibility through the evaluation of trans-eQTLs within GWAS loci. To this purpose, nine blood cell types and ileal, colonic and rectal biopsies have been collected for 330 healthy individuals of Northern European descent. All individuals have been genotyped with the OmniExpress Illumina array interrogating > 700K genetic variants. Transcriptome analysis has been conducted for all individuals and all cell/tissue types using Illumina HT12 arrays interrogating > 47,000 transcripts. Genotype and transcriptome data have undergone rigorous quality control. Transcriptome data have been pretreated variance stabilizing transformation, QQ normalization and correction for random and fixed effects in each cell type. Only expression probes mapped against Refseq have been considered. Genomic positions have been recovered and probes mapping to more than one genomic position (taking into account splice junctions with Tophat software) with a 96% identity have been discarded. Trans-eQTL mapping will be conducted on a SNP-by-SNP basis using linear regression (additive model) with PLINK software. In order to circumvent genome wide multiple testing penalty, we will test for a given SNP in the genome, any evidence for an excess of low p-values when testing its effect on the expression of genes located on other chromosomes or far away on the same chromosome. Confirmation of putative multigene transregulators will afterwards be performed by RNAseq experiments. We will then quantify the resemblance between DAP in the 160 GWAS-identified risk loci (raw data from IIBDGC plus imputed data) and " multigene trans-EAP " with Spearman's rank correlation. We will also evaluate the biological relevance of this list by performing a network analysis after adding the identified trans targets to the list of previously identified positional candidate genes (mapping to GWAS-identified IBD risk loci). Finally, as a the ultimate proof of causality, the selected genes will be resequenced in 3,000 IBD cases and 3,000 controls, using 600 DNA pools of 10 individuals with Illumina Truseq Amplicon. With this strategy, we expect to detect new causative variants that may constitute new drug targets for IBD. Latest results will be presented. [less ▲]

Genome Wide Association Studies (GWAS) have identified at least 68 loci involved in megakaryopoiesis and platelet formation. As for all GWAS, identified risk loci span hundreds of kilobases encompassing multiple genes, such that causative variants and genes remain largely unknown. To aid in the identification of causative genes underlying GWAS hits for platelet function (as well as other phenotypes including common complex diseases), we have generated a dataset (" CEDAR ") comprising genome-wide SNP and transcriptome data on nine primary cell types, including platelets, for 330 healthy Caucasian individuals. In addition, we have measured platelet counts and volume, as well as platelet reactivity to ADP, collagen and thrombin-related peptide for all these individuals. After extensive quality control, the ensuing data set has been used to identify (i) QTL influencing platelet count, volume and reactivity, and (ii) cis-and transacting eQTL operating in platelets. To aid in the identification of genes underlying platelet biology, we are applying a recently developed method to search for correlations between association patterns with platelet phenotypes and eQTL association patterns. Such findings would strongly incriminate the corresponding genes (affected by the eQTL) as being causally involved in determining the cognate platelet phenotype. Latest results will be presented. [less ▲]

Background: Research in epistasis or gene-gene interaction detection for human complex traits has grown over the last few years. It has been marked by promising methodological developments, improved ... [more ▼]

Background: Research in epistasis or gene-gene interaction detection for human complex traits has grown over the last few years. It has been marked by promising methodological developments, improved translation efforts of statistical epistasis to biological epistasis and attempts to integrate different omics information sources into the epistasis screening to enhance power. The quest for gene-gene interactions poses severe multiple-testing problems. In this context, the maxT algorithm is one technique to control the false-positive rate. However, the memory needed by this algorithm rises linearly with the amount of hypothesis tests. Gene-gene interaction studies will require a memory proportional to the squared number of SNPs. A genome-wide epistasis search would therefore require terabytes of memory. Hence, cache problems are likely to occur, increasing the computation time. In this work we present a new version of maxT, requiring an amount of memory independent from the number of genetic effects to be investigated. This algorithm was implemented in C++ in our epistasis screening software MBMDR-3.0.3. We evaluate the new implementation in terms of memory efficiency and speed using simulated data. The software is illustrated on real-life data for Crohn's disease. Results: In the case of a binary (affected/unaffected) trait, the parallel workflow of MBMDR-3.0.3 analyzes all gene-gene interactions with a dataset of 100,000 SNPs typed on 1000 individuals within 4 days and 9 hours, using 999 permutations of the trait to assess statistical significance, on a cluster composed of 10 blades, containing each four Quad-Core AMD Opteron Processor 2352 2.1 GHz. In the case of a continuous trait, a similar run takes 9 days. Our program found 14 SNP-SNP interactions with a multiple-testing corrected p-value of less than 0.05 on real-life Crohn's disease data. Conclusions: Our software is the first implementation of the MB-MDR methodology able to solve large-scale SNP-SNP interactions problems within a few days, without using much memory, while adequately controlling the type I error rates. A new implementation to reach genome-wide epistasis screening is under construction. In the context of Crohn's disease, MBMDR-3.0.3 could identify epistasis involving regions that are well known in the field and could be explained from a biological point of view. This demonstrates the power of our software to find relevant phenotype-genotype higher-order associations. [less ▲]

Epilepsy comprises several syndromes, amongst the most common being mesial temporal lobe epilepsy with hippocampal sclerosis. Seizures in mesial temporal lobe epilepsy with hippocampal sclerosis are ... [more ▼]

Epilepsy comprises several syndromes, amongst the most common being mesial temporal lobe epilepsy with hippocampal sclerosis. Seizures in mesial temporal lobe epilepsy with hippocampal sclerosis are typically drug-resistant, and mesial temporal lobe epilepsy with hippocampal sclerosis is frequently associated with important co-morbidities, mandating the search for better understanding and treatment. The cause of mesial temporal lobe epilepsy with hippocampal sclerosis is unknown, but there is an association with childhood febrile seizures. Several rarer epilepsies featuring febrile seizures are caused by mutations in SCN1A, which encodes a brain-expressed sodium channel subunit targeted by many anti-epileptic drugs. We undertook a genome-wide association study in 1018 people with mesial temporal lobe epilepsy with hippocampal sclerosis and 7552 control subjects, with validation in an independent sample set comprising 959 people with mesial temporal lobe epilepsy with hippocampal sclerosis and 3591 control subjects. To dissect out variants related to a history of febrile seizures, we tested cases with mesial temporal lobe epilepsy with hippocampal sclerosis with (overall n = 757) and without (overall n = 803) a history of febrile seizures. Meta-analysis revealed a genome-wide significant association for mesial temporal lobe epilepsy with hippocampal sclerosis with febrile seizures at the sodium channel gene cluster on chromosome 2q24.3 [rs7587026, within an intron of the SCN1A gene, P = 3.36 x 10(-9), odds ratio (A) = 1.42, 95% confidence interval: 1.26-1.59]. In a cohort of 172 individuals with febrile seizures, who did not develop epilepsy during prospective follow-up to age 13 years, and 6456 controls, no association was found for rs7587026 and febrile seizures. These findings suggest SCN1A involvement in a common epilepsy syndrome, give new direction to biological understanding of mesial temporal lobe epilepsy with hippocampal sclerosis with febrile seizures, and open avenues for investigation of prognostic factors and possible prevention of epilepsy in some children with febrile seizures. [less ▲]

Crohn's disease and ulcerative colitis, the two common forms of inflammatory bowel disease (IBD), affect over 2.5 million people of European ancestry, with rising prevalence in other populations. Genome ... [more ▼]

Crohn's disease and ulcerative colitis, the two common forms of inflammatory bowel disease (IBD), affect over 2.5 million people of European ancestry, with rising prevalence in other populations. Genome-wide association studies and subsequent meta-analyses of these two diseases as separate phenotypes have implicated previously unsuspected mechanisms, such as autophagy, in their pathogenesis and showed that some IBD loci are shared with other inflammatory diseases. Here we expand on the knowledge of relevant pathways by undertaking a meta-analysis of Crohn's disease and ulcerative colitis genome-wide association scans, followed by extensive validation of significant findings, with a combined total of more than 75,000 cases and controls. We identify 71 new associations, for a total of 163 IBD loci, that meet genome-wide significance thresholds. Most loci contribute to both phenotypes, and both directional (consistently favouring one allele over the course of human history) and balancing (favouring the retention of both alleles within populations) selection effects are evident. Many IBD loci are also implicated in other immune-mediated disorders, most notably with ankylosing spondylitis and psoriasis. We also observe considerable overlap between susceptibility loci for IBD and mycobacterial infection. Gene co-expression network analysis emphasizes this relationship, with pathways shared between host responses to mycobacteria and those predisposing to IBD. [less ▲]

In airways, the ecto-nucleoside triphosphate diphosphohydrolase CD39 plays a central role in the regulation of physiological mucosal nucleotide concentrations and likely contributes to the control of ... [more ▼]

In airways, the ecto-nucleoside triphosphate diphosphohydrolase CD39 plays a central role in the regulation of physiological mucosal nucleotide concentrations and likely contributes to the control of inflammation because accelerated ATP metabolism occurs in chronic inflammatory lung diseases.We sought to determine whether constant elevated CD39 activity in lung epithelia is sufficient to cause inflammation and whether this affects the response to acute LPS or Pseudomonas aeruginosa exposure. We generated transgenic mice overexpressing human CD39 under the control of the airway-specific Clara cell 10-kDa protein gene promoter. Transgenic mice did not develop any spontaneous lung inflammation. However, intratracheal instillation of LPS resulted in accelerated recruitment of neutrophils to the airways of transgenic mice. Macrophage clearance was delayed, and the amounts of CD8+ T and B cells were augmented. Increased levels of keratinocyte chemoattractant, IL-6, and RANTES were produced in transgenic lungs. Similarly, higher numbers of neutrophils and macrophages were found in the lungs of transgenic mice infected with P. aeruginosa, which correlated with improved bacteria clearance. The transgenic phenotype was partially and differentially restored by coinstillation of P2X1 or P2X7 receptor antagonists or of caffeine with LPS. Thus, a chronic increase of epithelial CD39 expression and activity promotes airway inflammation in response to bacterial challenge by enhancing P1 and P2 receptor activation. [less ▲]

Background: Research in epistasis or gene-gene interaction detection for human complex traits has grown exponentially over the last few years. It has been marked by promising methodological developments ... [more ▼]

Background: Research in epistasis or gene-gene interaction detection for human complex traits has grown exponentially over the last few years. It has been marked by promising methodological developments, improved translation efforts of statistical epistasis to biological epistasis and attempts to integrate different omics information sources into the epistasis screening to enhance power. The quest for gene-gene interactions poses severe multiple-testing problems. In this context, the maxT algorithm is one technique to control the false-positive rate. However, the memory needed by this algorithm rises linearly with the amount of hypothesis tests. In main-effects detection, this is not a problem since the memory required is thus proportional to the number of SNPs. In contrast, gene-gene interaction studies will require a memory proportional to the squared amount of SNPs. A genome wide epistasis would therefore require terabytes of memory. Hence, cache problems are likely to occur, increasing the computation time. Methods: In this work we present a new version of maxT, requiring an amount of memory independent from the number of genetic effects to be investigated. This algorithm was implemented in C++ in our epistasis screening software MB-MDR-2.6.2 and compared to MB-MDR's first implementation as an R-package (Calle et al., Bioinformatics 2010). We evaluate the new implementation in terms of memory efficiency and speed using simulated data. The software is illustrated on real-life data for Crohn's disease. Results: The sequential version of MBMDR-2.6.2 is approximately 5,500 times faster than its R counterparts. The parallel version (tested on a cluster composed of 14 blades, containing each 4 quad-cores Intel Xeon CPU E5520@2.27 GHz) is approximately 900,000 times faster than the latter, for results of the same quality on the simulated data. It analyses all gene-gene interactions of a dataset of 100,000 SNPs typed on 1000 individuals within 4 days. Our program found 14 SNP-SNP interactions with a p-value less than 0.05 on the real-life Crohn’s disease data. Conclusions: Our software is able to solve large-scale SNP-SNP interactions problems within a few days, without using much memory. A new implementation to reach genome wide epistasis screening is under construction. In the context of Crohn's disease, MBMDR-2.6.2 found signal in regions well known in the field and our results could be explained from a biological point of view. This demonstrates the power of our software to find relevant phenotype-genotype associations. [less ▲]

IL-1beta and TNF-alpha contribute to pancreatic beta cell death in type 1 diabetes. Both cytokines activate the transcription factor nuclear factor-kappaB (NF-kappaB), but recent observations suggest that NF-kappaB blockade prevents IL-1beta + IFN-gamma- but not TNF-alpha + IFN-gamma-induced beta cell apoptosis. The aim of the present study was to compare the effects of IL-1beta and TNF-alpha on cell death and the pattern of NF-kappaB activation and global gene expression in beta cells. METHODS: Cell viability was measured after exposure to IL-1beta or to TNF-alpha alone or in combination with IFN-gamma, and blockade of NF-kappaB activation or protein synthesis. INS-1E cells exposed to IL-1beta or TNF-alpha in time course experiments were used for IkappaB kinase (IKK) activation assay, detection of p65 NF-kappaB by immunocytochemistry, real-time RT-PCR and microarray analysis. RESULTS: Blocking NF-kappaB activation protected beta cells against IL-1beta + IFNgamma- or TNFalpha + IFNgamma-induced apoptosis. Blocking de novo protein synthesis did not increase TNF-alpha- or IL-1beta-induced beta cell death, in line with the observations that cytokines induced the expression of the anti-apoptotic genes A20, Iap-2 and Xiap to a similar extent. Microarray analysis of INS-1E cells treated with IL-1beta or TNF-alpha showed similar patterns of gene expression. IL-1beta, however, induced a higher rate of expression of NF-kappaB target genes putatively involved in beta cell dysfunction and death and a stronger activation of the IKK complex, leading to an earlier translocation of NF-kappaB to the nucleus. CONCLUSIONS/INTERPRETATION: NF-kappaB activation in beta cells has a pro-apoptotic role following exposure not only to IL-1beta but also to TNF-alpha. The more marked beta cell death induced by IL-1beta is explained at least in part by higher intensity NF-kappaB activation, leading to increased transcription of key target genes. [less ▲]